Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 2004 Mar 1;378(Pt 2):569–577. doi: 10.1042/BJ20031382

Anthrax lethal factor-cleavage products of MAPK (mitogen-activated protein kinase) kinases exhibit reduced binding to their cognate MAPKs.

A Jane Bardwell 1, Mahsa Abdollahi 1, Lee Bardwell 1
PMCID: PMC1223970  PMID: 14616089

Abstract

Anthrax lethal toxin is the major cause of death in systemic anthrax. Lethal toxin consists of two proteins: protective antigen and LF (lethal factor). Protective antigen binds to a cell-surface receptor and transports LF into the cytosol. LF is a metalloprotease that targets MKKs [MAPK (mitogen-activated protein kinase) kinases]/MEKs [MAPK/ERK (extracellular-signal-regulated kinase) kinases], cleaving them to remove a small N-terminal stretch but leaving the bulk of the protein, including the protein kinase domain, intact. LF-mediated cleavage of MEK1 and MKK6 has been shown to inhibit signalling through their cognate MAPK pathways. However, the precise mechanism by which this proteolytic cleavage inhibits signal transmission has been unclear. Here we show that the C-terminal LF-cleavage products of MEK1, MEK2, MKK3, MKK4, MKK6 and MKK7 are impaired in their ability to bind to their MAPK substrates, suggesting a common mechanism for the LF-induced inhibition of signalling.

Full Text

The Full Text of this article is available as a PDF (281.0 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Agrawal Anshu, Lingappa Jai, Leppla Stephen H., Agrawal Sudhanshu, Jabbar Abdul, Quinn Conrad, Pulendran Bali. Impairment of dendritic cells and adaptive immunity by anthrax lethal toxin. Nature. 2003 Jul 17;424(6946):329–334. doi: 10.1038/nature01794. [DOI] [PubMed] [Google Scholar]
  2. Bardwell A. J., Flatauer L. J., Matsukuma K., Thorner J., Bardwell L. A conserved docking site in MEKs mediates high-affinity binding to MAP kinases and cooperates with a scaffold protein to enhance signal transmission. J Biol Chem. 2000 Dec 28;276(13):10374–10386. doi: 10.1074/jbc.M010271200. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bardwell A. Jane, Abdollahi Mahsa, Bardwell Lee. Docking sites on mitogen-activated protein kinase (MAPK) kinases, MAPK phosphatases and the Elk-1 transcription factor compete for MAPK binding and are crucial for enzymic activity. Biochem J. 2003 Mar 15;370(Pt 3):1077–1085. doi: 10.1042/BJ20021806. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bardwell L., Cook J. G., Chang E. C., Cairns B. R., Thorner J. Signaling in the yeast pheromone response pathway: specific and high-affinity interaction of the mitogen-activated protein (MAP) kinases Kss1 and Fus3 with the upstream MAP kinase kinase Ste7. Mol Cell Biol. 1996 Jul;16(7):3637–3650. doi: 10.1128/mcb.16.7.3637. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bradley K. A., Mogridge J., Mourez M., Collier R. J., Young J. A. Identification of the cellular receptor for anthrax toxin. Nature. 2001 Nov 8;414(6860):225–229. doi: 10.1038/n35101999. [DOI] [PubMed] [Google Scholar]
  6. Bragg T. S., Robertson D. L. Nucleotide sequence and analysis of the lethal factor gene (lef) from Bacillus anthracis. Gene. 1989 Sep 1;81(1):45–54. doi: 10.1016/0378-1119(89)90335-1. [DOI] [PubMed] [Google Scholar]
  7. Chang Chung I., Xu Bing-e, Akella Radha, Cobb Melanie H., Goldsmith Elizabeth J. Crystal structures of MAP kinase p38 complexed to the docking sites on its nuclear substrate MEF2A and activator MKK3b. Mol Cell. 2002 Jun;9(6):1241–1249. doi: 10.1016/s1097-2765(02)00525-7. [DOI] [PubMed] [Google Scholar]
  8. Chang L., Karin M. Mammalian MAP kinase signalling cascades. Nature. 2001 Mar 1;410(6824):37–40. doi: 10.1038/35065000. [DOI] [PubMed] [Google Scholar]
  9. Charest D. L., Mordret G., Harder K. W., Jirik F., Pelech S. L. Molecular cloning, expression, and characterization of the human mitogen-activated protein kinase p44erk1. Mol Cell Biol. 1993 Aug;13(8):4679–4690. doi: 10.1128/mcb.13.8.4679. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Chopra Arun P., Boone Sherrie A., Liang Xudong, Duesbery Nicholas S. Anthrax lethal factor proteolysis and inactivation of MAPK kinase. J Biol Chem. 2003 Jan 9;278(11):9402–9406. doi: 10.1074/jbc.M211262200. [DOI] [PubMed] [Google Scholar]
  11. Davis R. J. Signal transduction by the JNK group of MAP kinases. Cell. 2000 Oct 13;103(2):239–252. doi: 10.1016/s0092-8674(00)00116-1. [DOI] [PubMed] [Google Scholar]
  12. Dixon T. C., Meselson M., Guillemin J., Hanna P. C. Anthrax. N Engl J Med. 1999 Sep 9;341(11):815–826. doi: 10.1056/NEJM199909093411107. [DOI] [PubMed] [Google Scholar]
  13. Dong Chen, Davis Roger J., Flavell Richard A. MAP kinases in the immune response. Annu Rev Immunol. 2001 Oct 4;20:55–72. doi: 10.1146/annurev.immunol.20.091301.131133. [DOI] [PubMed] [Google Scholar]
  14. Duesbery N. S., Resau J., Webb C. P., Koochekpour S., Koo H. M., Leppla S. H., Vande Woude G. F. Suppression of ras-mediated transformation and inhibition of tumor growth and angiogenesis by anthrax lethal factor, a proteolytic inhibitor of multiple MEK pathways. Proc Natl Acad Sci U S A. 2001 Mar 20;98(7):4089–4094. doi: 10.1073/pnas.061031898. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Duesbery N. S., Webb C. P., Leppla S. H., Gordon V. M., Klimpel K. R., Copeland T. D., Ahn N. G., Oskarsson M. K., Fukasawa K., Paull K. D. Proteolytic inactivation of MAP-kinase-kinase by anthrax lethal factor. Science. 1998 May 1;280(5364):734–737. doi: 10.1126/science.280.5364.734. [DOI] [PubMed] [Google Scholar]
  16. Enslen H., Brancho D. M., Davis R. J. Molecular determinants that mediate selective activation of p38 MAP kinase isoforms. EMBO J. 2000 Mar 15;19(6):1301–1311. doi: 10.1093/emboj/19.6.1301. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Enslen H., Davis R. J. Regulation of MAP kinases by docking domains. Biol Cell. 2001 Sep;93(1-2):5–14. doi: 10.1016/s0248-4900(01)01156-x. [DOI] [PubMed] [Google Scholar]
  18. Ezzell J. W., Ivins B. E., Leppla S. H. Immunoelectrophoretic analysis, toxicity, and kinetics of in vitro production of the protective antigen and lethal factor components of Bacillus anthracis toxin. Infect Immun. 1984 Sep;45(3):761–767. doi: 10.1128/iai.45.3.761-767.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Friedlander A. M. Tackling anthrax. Nature. 2001 Nov 8;414(6860):160–161. doi: 10.1038/35102660. [DOI] [PubMed] [Google Scholar]
  20. Guidi-Rontani Chantal. The alveolar macrophage: the Trojan horse of Bacillus anthracis. Trends Microbiol. 2002 Sep;10(9):405–409. doi: 10.1016/s0966-842x(02)02422-8. [DOI] [PubMed] [Google Scholar]
  21. Hale K. K., Trollinger D., Rihanek M., Manthey C. L. Differential expression and activation of p38 mitogen-activated protein kinase alpha, beta, gamma, and delta in inflammatory cell lineages. J Immunol. 1999 Apr 1;162(7):4246–4252. [PubMed] [Google Scholar]
  22. Hanna P. C., Ireland J. A. Understanding Bacillus anthracis pathogenesis. Trends Microbiol. 1999 May;7(5):180–182. doi: 10.1016/s0966-842x(99)01507-3. [DOI] [PubMed] [Google Scholar]
  23. Ho David T., Bardwell A. Jane, Abdollahi Mahsa, Bardwell Lee. A docking site in MKK4 mediates high affinity binding to JNK MAPKs and competes with similar docking sites in JNK substrates. J Biol Chem. 2003 Jun 3;278(35):32662–32672. doi: 10.1074/jbc.M304229200. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Johnson Gary L., Lapadat Razvan. Mitogen-activated protein kinase pathways mediated by ERK, JNK, and p38 protein kinases. Science. 2002 Dec 6;298(5600):1911–1912. doi: 10.1126/science.1072682. [DOI] [PubMed] [Google Scholar]
  25. Klimpel K. R., Arora N., Leppla S. H. Anthrax toxin lethal factor contains a zinc metalloprotease consensus sequence which is required for lethal toxin activity. Mol Microbiol. 1994 Sep;13(6):1093–1100. doi: 10.1111/j.1365-2958.1994.tb00500.x. [DOI] [PubMed] [Google Scholar]
  26. Kochi S. K., Schiavo G., Mock M., Montecucco C. Zinc content of the Bacillus anthracis lethal factor. FEMS Microbiol Lett. 1994 Dec 15;124(3):343–348. doi: 10.1111/j.1574-6968.1994.tb07306.x. [DOI] [PubMed] [Google Scholar]
  27. Koo Han-Mo, VanBrocklin Matt, McWilliams Mary Jane, Leppla Stephan H., Duesbery Nicholas S., Vande Woude George F. Apoptosis and melanogenesis in human melanoma cells induced by anthrax lethal factor inactivation of mitogen-activated protein kinase kinase. Proc Natl Acad Sci U S A. 2002 Feb 26;99(5):3052–3057. doi: 10.1073/pnas.052707699. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Lennon G., Auffray C., Polymeropoulos M., Soares M. B. The I.M.A.G.E. Consortium: an integrated molecular analysis of genomes and their expression. Genomics. 1996 Apr 1;33(1):151–152. doi: 10.1006/geno.1996.0177. [DOI] [PubMed] [Google Scholar]
  29. Liu S., Netzel-Arnett S., Birkedal-Hansen H., Leppla S. H. Tumor cell-selective cytotoxicity of matrix metalloproteinase-activated anthrax toxin. Cancer Res. 2000 Nov 1;60(21):6061–6067. [PubMed] [Google Scholar]
  30. Liu Shihui, Schubert Rebecca L., Bugge Thomas H., Leppla Stephen H. Anthrax toxin: structures, functions and tumour targeting. Expert Opin Biol Ther. 2003 Aug;3(5):843–853. doi: 10.1517/14712598.3.5.843. [DOI] [PubMed] [Google Scholar]
  31. Manning G., Whyte D. B., Martinez R., Hunter T., Sudarsanam S. The protein kinase complement of the human genome. Science. 2002 Dec 6;298(5600):1912–1934. doi: 10.1126/science.1075762. [DOI] [PubMed] [Google Scholar]
  32. Pannifer A. D., Wong T. Y., Schwarzenbacher R., Renatus M., Petosa C., Bienkowska J., Lacy D. B., Collier R. J., Park S., Leppla S. H. Crystal structure of the anthrax lethal factor. Nature. 2001 Nov 8;414(6860):229–233. doi: 10.1038/n35101998. [DOI] [PubMed] [Google Scholar]
  33. Park Jin Mo, Greten Florian R., Li Zhi-Wei, Karin Michael. Macrophage apoptosis by anthrax lethal factor through p38 MAP kinase inhibition. Science. 2002 Aug 29;297(5589):2048–2051. doi: 10.1126/science.1073163. [DOI] [PubMed] [Google Scholar]
  34. Pellizzari R., Guidi-Rontani C., Vitale G., Mock M., Montecucco C. Anthrax lethal factor cleaves MKK3 in macrophages and inhibits the LPS/IFNgamma-induced release of NO and TNFalpha. FEBS Lett. 1999 Nov 26;462(1-2):199–204. doi: 10.1016/s0014-5793(99)01502-1. [DOI] [PubMed] [Google Scholar]
  35. Popov Serguei G., Villasmil Rafael, Bernardi Jessica, Grene Edith, Cardwell Jennifer, Popova Taissia, Wu Aiguo, Alibek Darya, Bailey Charles, Alibek Ken. Effect of Bacillus anthracis lethal toxin on human peripheral blood mononuclear cells. FEBS Lett. 2002 Sep 11;527(1-3):211–215. doi: 10.1016/s0014-5793(02)03228-3. [DOI] [PubMed] [Google Scholar]
  36. Popov Serguei G., Villasmil Rafael, Bernardi Jessica, Grene Edith, Cardwell Jennifer, Wu Aiguo, Alibek Darya, Bailey Charles, Alibek Ken. Lethal toxin of Bacillus anthracis causes apoptosis of macrophages. Biochem Biophys Res Commun. 2002 Apr 26;293(1):349–355. doi: 10.1016/S0006-291X(02)00227-9. [DOI] [PubMed] [Google Scholar]
  37. Sharrocks A. D., Yang S. H., Galanis A. Docking domains and substrate-specificity determination for MAP kinases. Trends Biochem Sci. 2000 Sep;25(9):448–453. doi: 10.1016/s0968-0004(00)01627-3. [DOI] [PubMed] [Google Scholar]
  38. Struhl K. Reverse biochemistry: methods and applications for synthesizing yeast proteins in vitro. Methods Enzymol. 1991;194:520–535. doi: 10.1016/0076-6879(91)94039-f. [DOI] [PubMed] [Google Scholar]
  39. Tanoue T., Adachi M., Moriguchi T., Nishida E. A conserved docking motif in MAP kinases common to substrates, activators and regulators. Nat Cell Biol. 2000 Feb;2(2):110–116. doi: 10.1038/35000065. [DOI] [PubMed] [Google Scholar]
  40. Tournier C., Whitmarsh A. J., Cavanagh J., Barrett T., Davis R. J. The MKK7 gene encodes a group of c-Jun NH2-terminal kinase kinases. Mol Cell Biol. 1999 Feb;19(2):1569–1581. doi: 10.1128/mcb.19.2.1569. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Vitale G., Bernardi L., Napolitani G., Mock M., Montecucco C. Susceptibility of mitogen-activated protein kinase kinase family members to proteolysis by anthrax lethal factor. Biochem J. 2000 Dec 15;352(Pt 3):739–745. [PMC free article] [PubMed] [Google Scholar]
  42. Vitale G., Pellizzari R., Recchi C., Napolitani G., Mock M., Montecucco C. Anthrax lethal factor cleaves the N-terminus of MAPKKs and induces tyrosine/threonine phosphorylation of MAPKs in cultured macrophages. Biochem Biophys Res Commun. 1998 Jul 30;248(3):706–711. doi: 10.1006/bbrc.1998.9040. [DOI] [PubMed] [Google Scholar]
  43. Xia Y., Wu Z., Su B., Murray B., Karin M. JNKK1 organizes a MAP kinase module through specific and sequential interactions with upstream and downstream components mediated by its amino-terminal extension. Genes Dev. 1998 Nov 1;12(21):3369–3381. doi: 10.1101/gad.12.21.3369. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Xu Be, Stippec S., Robinson F. L., Cobb M. H. Hydrophobic as well as charged residues in both MEK1 and ERK2 are important for their proper docking. J Biol Chem. 2001 May 14;276(28):26509–26515. doi: 10.1074/jbc.M102769200. [DOI] [PubMed] [Google Scholar]
  45. Zheng C. F., Guan K. L. Properties of MEKs, the kinases that phosphorylate and activate the extracellular signal-regulated kinases. J Biol Chem. 1993 Nov 15;268(32):23933–23939. [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES